Pulmonary stem cells, related methods and kits of parts

Abstract

Isolated pulmonary stem cells, the isolated pulmonary stem cells being slowly dividing Oct-4 expressing cells forming individual colonies and able to undergoing terminal differentiation into a mature phenotype in vitro; a method to identify stem cells of a tissue; a kit for the identification of stem cell of a tissue; a method for identifying an infective agent able to infect pulmonary cells; a method for the production of a virus able to infect pulmonary cells and method for identifying a compound interfering with infection of a cell by an infective agent.

1 . Isolated pulmonary stem cells, the isolated pulmonary stem cells being slowly dividing cells forming individual colonies in vitro, the pulmonary stem cells expressing the marker Oct-4, the pulmonary stem cells able to undergoing terminal differentiation into a mature phenotype.  
     
     
         2 . The isolated pulmonary stem cells of  claim 1 , wherein the pulmonary stem cells further express the markers SSEA-I and Sca-1.  
     
     
         3 . The isolated pulmonary stem cells of  claim 1 , wherein the pulmonary stem cells further express the markers SSEA-3 SSEA-4 and Sca-1.  
     
     
         4 . A method to identify stem cells of a tissue, the method comprising: 
 identifying slowly dividing Oct-4 expressing cells of the tissue,    the identified slowly dividing Oct-4 expressing cells able to form colonies in vitro, the identified slowly dividing Oct-4 expressing cells, upon isolation in primary culture, able to differentiate in a mature phenotype.    
     
     
         5 . The method of  claim 4 , wherein identifying slowly dividing Oct-4 expressing cells comprises 
 identifying the slowly dividing cells of the tissue; and    identifying the slowly dividing cells expressing the stem cell marker Oct-4.    
     
     
         6 . The method of  claim 4 , wherein identifying slowly dividing cells of the tissue is performed by identifying BrdU retaining cells of the tissue.  
     
     
         7 . The method of  claim 4 , wherein identifying the Oct-4 expressing cells of the tissue is performed by immunofluorescence, RT-PCR or quantitative PCR.  
     
     
         8 . A kit of parts for the identification of stem cell of a tissue, the kit of parts comprising 
 a first identifier for the identification of slowly dividing cells; and    a second identifier for the identification of the expression of the marker Oct-4,    the first and the second identifier to be used to identify slowly dividing Oct-4 expressing cells, the identified slowly dividing Oct-4 expressing cells able to form colonies in vitro and to differentiate in a mature phenotype, being stem cells.    
     
     
         9 . The kit of parts of  claim 8 , wherein the first identifier is BrdU and the second identifier is selected from the group consisting of primary and secondary antibodies and oligonucleotides.  
     
     
         10 . The kit of parts of  claim 8 , wherein the tissue is a murine tissue and the kit further comprises a third identifier for the identification of an additional marker, the additional marker selected from the group consisting of SSEA-1 and SCA-1.  
     
     
         11 . The kit of parts of  claim 8 , wherein the tissue is a human tissue and the kit further comprises a third identifier for the identification of an additional marker, the additional marker selected from the group consisting of SSEA-3, SEEA-4 and Sca-1.  
     
     
         12 . The kit of parts of  claim 8 , further comprising a fourth identifier for the identification of the mature phenotype.  
     
     
         13 . A method for cultivating adult stem cells in vitro, the method comprising: 
 providing a tissue comprising tissue cells;    isolating the tissue cells from the tissue;    incubating the isolated tissue cells in a suitable medium;    applying to the isolated incubated cell a cell sorting procedure to obtain colonies in primary cultures; and    selecting the colonies in primary colonies that express a marker Oct-4, the colonies able to differentiate into mature type.    
     
     
         14 . The method of  claim 13 , wherein the adult stem cells are pulmonary stem cells.  
     
     
         15 . A method for identifying an infective agent able to infect pulmonary cells, the method comprising 
 contacting an infective agent with an isolated pulmonary stem cell;    detecting the level of infection of the isolated pulmonary stem cell by the infective agent; and    comparing the detected level of infection with a predetermined threshold level, the threshold level indicative of development of infection of the cell.    
     
     
         16 . A method for the production of a virus able to infect pulmonary cells, the method comprising: 
 contacting a virus particle with an isolated pulmonary stem cell in a pulmonary stem cell culture; and    collecting virus particles from the cell culture.    
     
     
         17 . The method of  claim 16 , wherein contacting a virus particle with an isolated pulmonary stem cell in a culture is able be performed in a serum-free culture condition.  
     
     
         18 . The method of  claim 17 , wherein the viral particle is selected from the group consisting of influenza and avian flu viral particles.  
     
     
         19 . The method of  claim 16 , wherein the viral particle is a SARS viral particle.  
     
     
         20 . A method for identifying a compound interfering with infection of a pulmonary stem cell by an infective agent, the method comprising 
 contacting a candidate compound with an isolated pulmonary stem cell;    contacting the isolated pulmonary stem cell with a candidate infective agent;    detecting the level of infection of the isolated pulmonary stem cell by the infective agent; and    comparing the detected level of infection with a predetermined threshold level, the threshold level indicative of development of infection of the cell.    
     
     
         21 . The method of  claim 20 , wherein the infective agent is a virus, the virus selected from the group consisting of influenza and avian flues viruses.  
     
     
         22 . The method of  claim 20 , wherein the infective agent is SARS virus.